RESUMEN
Staphylococcus aureus is a serious threat to public health due to the rise of antibiotic resistance in this organism, which can prolong or exacerbate skin and soft tissue infections (SSTIs). Methicillin-resistant S. aureus is a Gram-positive bacterium and a leading cause of SSTIs. As such, many efforts are under way to develop therapies that target essential biological processes in S. aureus. Antimicrobial photodynamic therapy is an effective alternative to antibiotics; therefore we developed an approach to simultaneously expose S. aureus to intracellular and extracellular photosensitizers. A near infrared photosensitizer was conjugated to human monoclonal antibodies (MAbs) that target the S. aureus iron-regulated surface determinant (Isd) heme acquisition proteins. In addition, the compound VU0038882 was developed to increase photoactivatable porphyrins within the cell. Combinatorial photodynamic treatment of drug-resistant S. aureus exposed to VU0038882 and conjugated anti-Isd MAbs proved to be an effective antibacterial strategy in vitro and in a murine model of SSTIs.
Asunto(s)
Staphylococcus aureus Resistente a Meticilina , Infecciones de los Tejidos Blandos , Infecciones Estafilocócicas , Animales , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Humanos , Ratones , Fármacos Fotosensibilizantes/farmacología , Infecciones de los Tejidos Blandos/tratamiento farmacológico , Infecciones Estafilocócicas/tratamiento farmacológico , Staphylococcus aureusRESUMEN
A fluorescently labeled, persulfated molecular umbrella ( 1) has been synthesized from cholic acid, lysine, spermine, and Coumarin 343 and found capable of entering live HeLa cells. The distributions of 1 throughout the cytoplasm and the nucleus were diffuse and punctate, respectively. This finding, together with its ability to cross liposomal membranes by passive diffusion, suggests that passive diffusion plays a significant role in the ability of 1 to enter cells. The fact that 1 is concentrated at the nucleus raises the possibility that molecular umbrellas of this type could be used for the nuclear targeting of drugs.
Asunto(s)
Núcleo Celular/metabolismo , Portadores de Fármacos/química , Portadores de Fármacos/metabolismo , Aniones/química , Membrana Celular/metabolismo , Colorantes Fluorescentes/química , Células HeLa , Humanos , Membrana Dobles de Lípidos/metabolismo , Liposomas/metabolismo , Microscopía Confocal , Péptidos/metabolismo , Agua/químicaRESUMEN
One of the most challenging questions that relates to the structure and function of biological membranes is whether the two halves of the bilayer "talk" to each other. In this letter, we show how the perturbation of the lateral organization of one leaflet of a fluid phospholipid bilayer by an external agent also alters the lateral organization of the adjoining leaflet. In addition, we show that the energy involved in such "cross talk" corresponds to ca. 100 cal/mol of phospholipid. These findings provide a basis for expecting similar cross talk to exist in cell membranes.
Asunto(s)
Membrana Dobles de Lípidos/química , Fosfolípidos/química , Dimerización , Estructura Molecular , Proyectos de InvestigaciónRESUMEN
Molecular umbrella compounds may function as novel topical microbicides to prevent human immunodeficiency virus (HIV) and herpes simplex virus (HSV) infections. In a preliminary structure-activity investigation, one umbrella compound, designated Spm8CHAS, was identified which inhibited both HIV and HSV infections with no cellular toxicity. The objectives of the current studies were to define its spectrum of antiviral activity, characterize its mechanism of action, and explore the possibility of combining Spm8CHAS with HIV-specific reverse transcriptase inhibitors. Spm8CHAS inhibited infections by laboratory and clinical R5 and X4 clade B and clade C HIV strains in cell culture. Ectocervical tissue explants exposed to HIV-1(BaL) in the presence of Spm8CHAS were completely protected (50% inhibitory concentration [IC(50)], 13.6 microg/ml), and transfer of virus to target T cells via migratory cells was abolished (IC(50), 3.8 microg/ml). Spm8CHAS inhibited HSV-2 infection of epithelial cells 10,000-fold if present throughout the infection. Notably, adding Spm8CHAS to cultures following HSV entry significantly reduced viral infection, indicating that the drug also acts postentry. Subsequent studies indicated that Spm8CHAS blocks cell-to-cell spread of HSV. Confocal microscopy using a fluorescently labeled analog of Spm8CHAS demonstrated that this conjugate crosses the plasma cell membrane and is transported to the nucleus. Combinations of Spm8CHAS with UC-781 or 9-[R-2-(phosphonylmethoxy)propyl] adenine monohydrate in vitro exhibited additive anti-HIV activity with preserved anti-HSV activity. The abilities of Spm8CHAS to inhibit primary isolates of HIV, block HSV infection postentry, and cross cell membranes support the development of a combination microbicide containing Spm8CHAS with an HIV-specific reverse transcriptase inhibitor to prevent both HIV and HSV infections by multiple mechanisms.
Asunto(s)
Antivirales/uso terapéutico , Infecciones por VIH/prevención & control , Herpes Simple/prevención & control , Células Cultivadas , HumanosRESUMEN
The mixing behavior of exchangeable, disulfide-based mimics of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and cholesterol has been examined as a function of temperature in host membranes made from DPPC and cholesterol in the liquid-disordered phase (ld), in the liquid-ordered phase (lo), and in the liquid-disordered/liquid-ordered coexistence region (ld/lo). In the ld region, lipid mixing was found to be temperature insensitive, reflecting close to ideal behavior. In contrast, a significant temperature dependence was observed in the lo phase from 45 to 60 degrees C, when 35 or 40 mol % sterol was present. In this region, sterol-phospholipid association was characterized by DeltaHo = -2.06 +/- 0.14 kcal/mol of phospholipid and DeltaS degrees = -4.48 +/- 0.44 cal/K mol of phospholipid. From 60 to 65 degrees C, the mixing of these lipids was found to be insensitive to temperature, and sterol-phospholipid association was now entropy driven; that is, DeltaHo = -0.23 +/- 0.38 kcal/mol of phospholipid and DeltaS degrees = +1.68 +/- 1.12 cal/K mol of phospholipid. In the liquid-disordered/liquid-ordered coexistence region, changes in lipid mixing reflect changes in the phase composition of the membrane.
Asunto(s)
1,2-Dipalmitoilfosfatidilcolina/química , Colesterol/química , Membrana Dobles de Lípidos/química , Fluidez de la Membrana , Modelos Químicos , Modelos Moleculares , 1,2-Dipalmitoilfosfatidilcolina/análisis , Colesterol/análisis , Mezclas Complejas/análisis , Mezclas Complejas/química , Simulación por Computador , Membrana Dobles de Lípidos/análisis , Sustancias Macromoleculares/análisis , Sustancias Macromoleculares/química , Transición de Fase , TermodinámicaRESUMEN
This paper records what is believed to be the first evidence for the reorganization of the liquid-ordered phase by ethanol. Specifically, ethanol has been found to significantly enhance sterol-phospholipid association in liquid-ordered bilayers derived from 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) plus cholesterol and also 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) plus cholesterol. The evidence for such reorganization comes from a series of nearest-neighbor recognition (NNR) experiments that have been carried out, where low concentrations of equilibrating lipid dimers (i.e., "reporter molecules") have been used to detect changes in the phase composition of host membranes made from varying mixtures of DPPC/cholesterol, and also DSPC/cholesterol, in the presence and in the absence of ethanol. These findings have important biological implications, which are briefly discussed.
Asunto(s)
1,2-Dipalmitoilfosfatidilcolina/química , Colesterol/química , Etanol/química , Membrana Dobles de Lípidos/química , Fosfatidilcolinas/química , Etanol/farmacología , CinéticaRESUMEN
A series of molecular umbrella conjugates, derived from cholic acid, deoxycholic acid, spermidine, lysine, and 5-mercapto-2-nitrobenzoic acid, have been synthesized and found capable of transporting an attached 16-mer oligonucleotide (S-dT16) across liposomal membranes made from 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidyldglycerol (POPG), and cholesterol [POPC/POPG/cholesterol (65/5/30; mol/mol/mol, v/v/v)] at 37 degrees C. Those molecular umbrellas containing four choloyl (or deoxycholoyl) groups resulted in significantly faster rates of transport as compared to those containing only two such moieties. A model that accounts for these membrane transport processes is proposed.
Asunto(s)
Colesterol/química , Membrana Dobles de Lípidos/química , Oligonucleótidos/metabolismo , Fosfolípidos/química , Transporte Biológico , Ácido Cólico/química , Ácido Desoxicólico/química , Membrana Dobles de Lípidos/síntesis química , Liposomas/química , Lisina/química , Modelos Moleculares , Nitrobenzoatos/química , Oligonucleótidos/química , Fosfatidilcolinas/química , Fosfatidilgliceroles/química , Espermidina/química , Relación Estructura-Actividad , Compuestos de SulfhidriloRESUMEN
A series of nearest-neighbor recognition (NNR) experiments have been carried out, which provide a rigorous test of the existence of transbilayer complementarity of phospholipids, that is, the ability of phospholipids to select complementary phospholipids from an adjoining monolayer as nearest neighbors. The application of this test to membranes derived from exchangeable phospholipids bearing myristoyl groups (A), stearoyl groups (B), and one stearoyl and one n-dodecyl group (C) in the presence of analogous nonexchangeable templates made from A', B' and C' provides compelling evidence for such complementarity in the physiologically relevant fluid phase.
Asunto(s)
Membrana Dobles de Lípidos/química , Fluidez de la Membrana/fisiología , Fosfolípidos/química , Cromatografía Líquida de Alta Presión , Interacciones Hidrofóbicas e Hidrofílicas , Membrana Dobles de Lípidos/metabolismo , Microdominios de Membrana/química , Microdominios de Membrana/metabolismo , Modelos Químicos , Miristatos/química , Miristatos/metabolismo , Fosfolípidos/metabolismo , Ácidos Esteáricos/química , Ácidos Esteáricos/metabolismoRESUMEN
The mixing properties of exchangeable phospholipids, derived from 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine, with an exchangeable form of cholesterol have been used to monitor the transition from the liquid-disordered to the liquid-ordered phase in cholesterol-containing bilayers, made from 1,2-dipalmitoyl-sn-glycero-3-phosphocholine and 1,2-distearoyl-sn-glycero-3-phosphocholine, respectively.
Asunto(s)
Membrana Dobles de Lípidos/química , Lípidos de la Membrana/química , 1,2-Dipalmitoilfosfatidilcolina/química , Colesterol/química , Cinética , Fosfatidilcolinas/química , Fosfatidiletanolaminas/químicaRESUMEN
Lipid-lipid interactions across cholesterol-rich phospholipid bilayers were investigated by measuring nearest-neighbor preferences of exchangeable phospholipids derived from 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine (DMPE) and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine (DSPE), in the presence of nonexchangeable dimers (i.e., templates) made from DMPE or DSPE. When homotemplates were present, a significant preference for homophospholipid association was observed. In contrast, when the corresponding heterotemplate was present, heterodimer formation was favored. These results support a model in which the longer phospholipid in one monolayer preferentially associates with the shorter one in the adjoining monolayer. In the absence of cholesterol, transbilayer complementarity was also observed but to a lesser degree. Transbilayer complementarity of phospholipids is likely to play an important role in stabilizing biological membranes and in promoting a compositional interdependence of their two lipid leaflets.
Asunto(s)
Colesterol/química , Membrana Dobles de Lípidos/química , Modelos Químicos , Fosfolípidos/química , Colesterol/metabolismo , Dimerización , Disulfuros/química , Interacciones Hidrofóbicas e Hidrofílicas , Liposomas , Microdominios de Membrana/química , Fosfatidiletanolaminas/química , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Compuestos de Sulfhidrilo/químicaRESUMEN
A series of persulfated molecular umbrellas have been synthesized from putrescine, spermidine, spermine, lysine, and cholic acid (1a, 2a, 3a, 4a, and 5a) and their anti-HIV and anti-HSV activities determined. Despite it size, the most active of these conjugates (5a) was able to cross phospholipid bilayers made from 1-palmitoyl-2-oleyol-sn-glycero-3-phosphocholine (POPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylglycerol (POPG). The unique combination of anti-viral activity, an ability to cross hydrophobic barriers, a lack of cytotoxicity, and a simple three-step synthesis from biogenic starting material suggests that 5a and related conjugates may be exploitable as a novel class of anti-viral agents for systemic and topical applications.
Asunto(s)
Antivirales/química , Antivirales/farmacología , Ácidos Cólicos/química , Ácidos Cólicos/farmacología , VIH-1/efectos de los fármacos , Herpesvirus Humano 2/efectos de los fármacos , Ésteres del Ácido Sulfúrico/química , Ésteres del Ácido Sulfúrico/farmacología , Fármacos Anti-VIH/administración & dosificación , Fármacos Anti-VIH/química , Fármacos Anti-VIH/farmacocinética , Fármacos Anti-VIH/farmacología , Antivirales/administración & dosificación , Antivirales/farmacocinética , Línea Celular , Membrana Celular/metabolismo , Cuello del Útero/virología , Femenino , Herpesvirus Humano 2/metabolismo , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Membrana Dobles de Lípidos/química , Membrana Dobles de Lípidos/metabolismo , Relación Estructura-ActividadRESUMEN
The mixing behavior of a phospholipid containing a cis-cyclopropyl moiety (1) with one that contains two myristoyl groups (3a) has been investigated in fluid bilayers via the nearest-neighbor recognition (NNR) method. In the absence of cholesterol, these lipids mix ideally. In the presence of cholesterol, they show a modest preference for homo-phospholipid association. A trans-form of 1 (i.e., 2) was found to have similar behavior, except that the influence of cholesterol in promoting homo-phospholipid association was greater. Similar results have been found in membranes in which 3a is replaced with a phospholipid bearing two palmitoyol chains (3b). In this case, the effect of the kink is approximately twice as great. The implications of these findings, with respect to the "trans-fatty acid debate", are briefly discussed
Asunto(s)
Colesterol/química , Membrana Dobles de Lípidos/química , Fosfolípidos/química , Conformación Molecular , TermodinámicaRESUMEN
Nearest-neighbor recognition experiments, which have been carried out using exchangeable dimers derived from 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine, 1,2-distearoyl-sn-glycero-3-phosphoethanolamine,and 1-palmitoyl-2-oleoyl-sn-glycerophosphoethanolamine, indicate that replacement of H2O by D2O can significantly influence phospholipid mixing, but only in bilayers that are saturated and devoid of cholesterol. These findings, together with those of previous electron spin resonance spin-labeling studies,indicate that mammalian membranes, which are rich in cholesterol and unsaturated phospholipids, are ideal hydrophobic barriers.
Asunto(s)
Membrana Dobles de Lípidos/química , Fosfolípidos/química , Fenómenos Químicos , Química Física , Colesterol/química , Deuterio/química , Medición de Intercambio de Deuterio , Estructura Molecular , Solventes/química , Propiedades de Superficie , Temperatura , Agua/químicaRESUMEN
Lipid-lipid interactions across a phospholipid bilayer were probed by measuring the nearest-neighbor preferences of exchangeable phospholipid monomers derived from 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine (DMPE) and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine (DSPE) in the presence of nonexchangeable DMPE- or DSPE-based dimers. Each of these permanent dimers promoted homophospholipid association to the same extent, whereas the corresponding nonexchangeable monomers were without effect. These results support a model in which the longer phospholipids in one monolayer leaflet preferentially associate with shorter ones in the adjoining monolayer. Such transbilayer complementarity is likely to play an important role in stabilizing biological membranes and also in promoting a compositional interdependence of their two lipid leaflets.
Asunto(s)
Membrana Dobles de Lípidos/química , Membrana Dobles de Lípidos/metabolismo , Fosfolípidos/química , Fosfolípidos/metabolismo , Cinética , Modelos Biológicos , Modelos MolecularesRESUMEN
Three Ru(II) complexes of type as [Ru(II)(bpy)(2)L](2+) were synthesized, where L are l,10-phenanthroline derivatives of imidazole (1), having at position 2 alpha-naphthyl (2), 3-methoxy-4-hydroxy-phenyl (3). All complexes show intense MLCT transition both in acetonitrile and in water and also exhibit strong emission at room temperature, which is efficiently quenched by oxygen as well as, to some extent, by water. The binding of complexes 1-3 to calf thymus DNA was investigated by using electronic absorption, steady-state luminescence, luminescence quenching, excited-state lifetime and circular dichroism spectra. Hypochromic effect, luminescence enhancement, and quenching studies demonstrate the existence of intercalation mode. Circular dichroism spectra indicate the stereoselectivity of the binding. The binding of 1-3 with DNA is sensitive to the nature of ligands, such as planarity, pi-electron extension and hydrophobicity. Complex 3 exhibits the strongest binding with DNA, which can be attributed to hydrogen bonding.
Asunto(s)
ADN/metabolismo , Compuestos Organometálicos/química , Fenantrolinas/química , Rutenio/química , Dicroismo Circular , Espectroscopía de Resonancia Magnética , Compuestos Organometálicos/metabolismo , Fenantrolinas/metabolismo , Rutenio/metabolismoRESUMEN
OBJECTIVE: To study the diagnostic effect of bronchoalveolar lavage in early lung injury by observing changes in inflammatory mediators in early lung injury caused by enterogenic infection. METHODS: Eighty-four Sprague-Dawley rats were randomly divided into infection group and sham-operation group. Cecal ligation and perforation was utilized to produce abdominal infection in rats. Six groups were sacrificed respectively at 0, 24, 48, 72, 96, 120 hours after operation. The differential cell count in bronchoalveolar lavage fluid (BALF) was assessed. The concentrations of endotoxin, phospholipase A2 (PLA2) and tumor necrosis factor-alpha (TNF-alpha) in BALF, lung and plasma were assayed. RESULTS: The neutrophil percentage of BALF increased progressively. The concentrations of endotoxin, PLA2 and TNF-alpha in BALF, lung and plasma were significantly increased. The levels of endotoxin and PLA2 in lung tissue were respectively correlated positively with those in BALF and plasma (BALF and lung: r=0.904, P<0.05; BALF and plasma: r=0.895, P<0.05; lung and plasma: r=0.946, P<0.01). Significant positive correlation was also present between the TNF-alpha levels in BALF and lung (r=0.952 P<0.01), but not between the TNF-alpha level in plasma and that in lung or BALF (r=0.684, r=0.608, both P>0.05). CONCLUSION: The examinations of bronchoalveolar lavage may help discover early lung injury caused by enterogenic infection.
Asunto(s)
Adenosina Monofosfato/análogos & derivados , Síndrome de Dificultad Respiratoria/diagnóstico , Adenosina Monofosfato/toxicidad , Animales , Líquido del Lavado Bronquioalveolar/química , Modelos Animales de Enfermedad , Endotoxinas/análisis , Femenino , Masculino , Oligopéptidos/toxicidad , Fosfolipasas A/análisis , Fosfolipasas A2 , Distribución Aleatoria , Ratas , Ratas Sprague-Dawley , Síndrome de Dificultad Respiratoria/inducido químicamente , Síndrome de Dificultad Respiratoria/metabolismo , Factor de Necrosis Tumoral alfa/análisisRESUMEN
Nearest-neighbor recognition measurements have established that the effects of deuterium substitution on phospholipid mixing are exceedingly small. Thus, the mixing behavior of an exchangeable phospholipid bearing two stearoyl chains with a homologue containing two myristoyl chains in gel-fluid bilayers, fluid bilayers, cholesterol-rich fluid bilayers, and gel-fluid bilayers that have been enriched with cholesterol correspond to a difference in the free energy of mixing that is less than 2.2 cal/mol of hydrogen in all cases. These findings provide the strongest evidence to date in support of the use of deuterated phospholipids as "nonperturbing" probes for structural and dynamic studies of bilayer membranes.